Glucocorticoids (GCs) are steroid hormones that regulate multiple physiological processes involved in inflammation, immunity, metabolism and homeostatic functions. They exert their effects by binding to the glucocorticoid receptor (GR, NR3C1), triggering its activation and translocation to the nucleus, leading to transcriptional changes responsible for diminished proliferative capacity and leukemia cell death (Yudt and Cidlowski, 2002).
Synthetic glucocorticoids (e.g., Hydrocortisone (Plenadren, Cortef), Prednisone, Dexamethasone (Intensol), Methylprednisolone (Medrol), Prednisolone (Orapred, Pediapred, Prelone)) are widely prescribed medications used to treat a variety of human diseases with an inflammatory component such as, e.g., asthma, allergies, chronic obstructive pulmonary disease (COPD), swelling, organ transplants, sarcoidosis, spinal cord injuries, ulcerative colitis, irritation, rheumatoid arthritis, chronic inflammatory demyelinating polyneuropathy, Addison's disease, multiple sclerosis (MS), and other autoimmune disorders. However, success of these glucocorticoid treatments is frequently hampered by resistance. For instance, glucocorticoid resistance in asthma has been estimated at 20%-50% (Barnes et al., Am. J. Respir. Critical Care Med., 1995, 152:5125-5142).
Glucocorticoids (e.g., prednisolone and dexamethasone) are also an essential component of curative therapy of acute lymphoblastic leukemia (ALL) and lymphomas. Advancements in the treatment of children with ALL have led to five-year disease-free survival rates exceeding 85% (Pui et al., 2009). However, children whose ALL cells show in vitro resistance to glucocorticoids have a significantly worse treatment outcome (disease-free survival) than patients whose ALL cells are sensitive to glucocorticoids (Den Boer et al., 2003; Dordelmann et al., 1999; Kaspers et al., 1997; Pieters et al., 1991). Yet, little is known about the mechanisms causing the leukemia cells from some patients to exhibit de novo resistance to glucocorticoids.
To find ways to overcome glucocorticoid resistance in acute lymphoblastic leukemia (ALL), lymphomas and other cancers is an urgent problem.
Caspase-1 (CASP1, also known as Interleukin-1β converting enzyme (ICE), apoptosis-related cysteine peptidase, IL-1β convertase, P45 and IL1B convertase, and IL1BC), is an intracellular protease that is known to cleave the precursors of IL-1β and IL-18 into active cytokines (Black et al., FEBS Lett, 247: 386-390 (1989); Kostura et al., Proc. Natl. Acad. Sci. U.S.A., 86:5227-5231 (1989)). Enzymatically active CASP1 is a heterotetramer composed of two subunits of p20 and two subunits of p10 (20 kDa and 10 kDa molecular weight, respectively). These subunits are derived from a 45 kDa proenzyme (p45) by way of a p30 form, through an activation mechanism that is autocatalytic (Thornberry et al., Nature, 356, pp. 768-774 (1992)). The CASP1 proenzyme has been divided into several functional domains: a prodomain (pi 4), a p22/20 subunit, a polypeptide linker and a p10 subunit (Thornberry et al., Nature, 356, pp. 768-774 (1992); Casano et al., Genomics, 20, pp. 474-481 (1994)).
CASP1 belongs to a family of cysteine proteases that cleave proteins following an aspartic acid residue. Produced as a pro-enzyme, CASP1 requires removal of its CARD (caspase activation and recruitment domain) before it becomes an active enzyme (Schroder and Tschopp, 2010). CARD cleavage is mediated by the formation of large complexes termed inflammasomes, of which the most extensively characterized is the NLRP3-containing inflammasome. NLRP3 (encoded by NLRP3 gene) can be activated by exposure to pathogen associated molecular pattern (PAMP) or damage associated molecular pattern (DAMP) molecules, or by whole pathogens or environmental irritants (Schroder and Tschopp, 2010). There is also emerging evidence that the NLRP3 inflammasome can form in response to host-derived molecules, including extracellular ATP, glucose or monosodium urate crystals (Mariathasan et al., 2006; Martinon et al., 2006; Schroder and Tschopp, 2010; Zhou et al., 2010). In vivo induction of the NLRP3 inflammasome typically results in self-oligomerization, recruitment of the ASC (PYCARD) adaptor protein, and clustering and autoactivation of CASP1.
Human CASP1 has the following sequence (SEQ ID NO: 1, corresponds to GenBank Accession No. NP 150634.1), wherein residues 1-119 of correspond to the propeptide region and residues 120-404 correspond to the mature chain. The p20 and p10 subunits correspond to residues 120-297 (p20) and residues 317-404 (p10), respectively.
(SEQ ID NO: 1)MADKVLKEKRKLFIRSMGEGTINGLLDELLQTRVLNKEEMEKVKRENA TVMDKTRALIDSVIPKGAQACQICITYICEEDSYLAGTLGLSADQTSG NYLNMQDSQGVLSSFPAPQAVQDNPAMPTSSGSEGNVKLCSLEEAQRI WKQKSAEIYPIMDKSSRTRLALIICNEEFDSIPRRTGAEVDITGMTML LQNLGYSVDVKKNLTASDMTTELEAFAHRPEHKTSDSTFLVFMSHGIR EGICGKKHSEQVPDILQLNAIFNMLNTKNCPSLKDKPKVIIIQACRGD SPGVVWFKDSVGVSGNLSLPTTEEFEDDAIKKAHIEKDFIAFCSSTPD NVSWRHPTMGSVFIGRLIEHMQEYACSCDVEEIFRKVRFSFEQPDGRA QMPTTERVTLTRCFYLFPGH
See also the following GenBank Accession Nos.:
Alpha precursor isoforms: NP_150634.1 (as above); NP_001244047.1
Beta precursor isoforms: NP_001214.1; NP_001244048.1
Gamma precursor isoforms: NP_150635.1
Delta precursor isoforms: NP_150636.1
Epsilon precursor isoforms: NP_150637.1
Human NLRP3 has the following sequence (SEQ ID NO: 2; corresponds to GenBank Accession No. NP_004886.3), wherein residues 8-93 of correspond to the pyrin death domain found in NALP proteins and residues 220-389 correspond to the NACHT domain and resides 575-891 correspond to Leucine-rich repeats (LRRs), ribonuclease inhibitor domain respectively.
(SEQ ID NO: 2)MKMASTRCKLARYLEDLEDVDLKKFKMHLEDYPPQKGCIPLPRGQTEK ADHVDLATLMIDFNGEEKAWAMAVWIFAAINRRDLYEKAKRDEPKWGS DNARVSNPTVICQEDSIEEEWMGLLEYLSRISICKMKKDYRKKYRKYV RSRFQCIEDRNARLGESVSLNKRYTRLRLIKEHRSQQEREQELLAIGK TKTCESPVSPIKMELLFDPDDEHSEPVHTVVFQGAAGIGKTILARKMM LDWASGTLYQDRFDYLFYIHCREVSLVTQRSLGDLIMSCCPDPNPPIH KIVRKPSRILFLMDGFDELQGAFDEHIGPLCTDWQKAERGDILLSSLI RKKLLPEASLLITTRPVALEKLQHLLDHPRHVEILGFSEAKRKEYFFK YFSDEAQARAAFSLIQENEVLFTMCFIPLVCWIVCTGLKQQMESGKSL AQTSKTTTAVYVFFLSSLLQPRGGSQEHGLCAHLWGLCSLAADGIWNQ KILFEESDLRNHGLQKADVSAFLRMNLFQKEVDCEKFYSFIHMTFQEF FAAMYYLLEEEKEGRTNVPGSRLKLPSRDVTVLLENYGKFEKGYLIFV VRFLFGLVNQERTSYLEKKLSCKISQQIRLELLKWIEVKAKAKKLQIQ PSQLELFYCLYEMQEEDFVQRAMDYFPKIEINLSTRMDHMVSSFCIEN CHRVESLSLGFLHNMPKEEEEEEKEGRHLDMVQCVLPSSSHAACSHGL VNSHLTSSFCRGLFSVLSTSQSLTELDLSDNSLGDPGMRVLCETLQHP GCNIRRLWLGRCGLSHECCFDISLVLSSNQKLVELDLSDNALGDFGIR LLCVGLKHLLCNLKKLWLVSCCLTSACCQDLASVLSTSHSLTRLYVGE NALGDSGVAILCEKAKNPQCNLQKLGLVNSGLTSVCCSALSSVLSTNQ NLTHLYLRGNTLGDKGIKLLCEGLLHPDCKLQVLELDNCNLTSHCCWD LSTLLTSSQSLRKLSLGNNDLGDLGVMMFCEVLKQQSCLLQNLGLSEM YFNYETKSALETLQEEKPELTVVFEPSW
See also the following GenBank Accession Nos.:
Isoform a: NP_004886.3 (as above); NP_001073289.1
Isoform b: NP_899632.1
Isoform c: NP_001120933.1
Isoform d: NP_001120934.1
Isoform e: NP_001230062.1